Research news

Researchers based in the United States have discovered significant differences in
the regulation of gene splicing between mammals and fish. Their findings, reported in Proceedings of the National Academy of Sciences this week, could help scientists develop transgenic techniques using pufferfish DNA
sequences in mouse and human cells (Proc Natl Acad Sci USA 2004, 101:15700-15705).

The genome of the pufferfish - or Fugu - contains all the alternative promoters and
splice exons and introns that are present in mammalian genomes, but because the introns are so much smaller,
genes are about an eighth the size, said lead author Christopher B. Burge, at Massachusetts Institute of Technology (MIT).

This makes the Fugu genome a potentially powerful tool for functional gene analysis,
Burge said, but scientists have until now been frustrated in their attempts to use
the resource because mammalian cells do not correctly splice the fish genes.

Burge's team developed a variant of a previously devised method for predicting splicing enhancer sequences. The new technique - dubbed RESCUE-ISE
- predicts intronic splicing enhancers (ISEs), and by comparing human, mouse, zebrafish,
and Fugu genomes, Burge's group discovered that this class of splicing regulatory
element appears to differ substantially between mammals and fish.

Burge told The Scientist that "89 to 96% of all the hexamers that we predicted as ISEs in mammals fall into
one of two clusters - they're either G-rich or C-rich. But when we applied the same
method to Fugu introns, we just got a completely different spectrum of motifs."

Burge proposes that by applying a scoring method to individual intron sequences, Fugu
genes can be tested for problem motifs and modified for transgenic experiments in
mice - a method he has successfully piloted. "You have to do a bit of extra cloning
or site-directed mutagenesis," he said. "But these Fugu genes would be much easier
to manipulate and that much more genetically tractable."

"I think the paper's of interest in terms of understanding the potential of regulation
and some of the details of the mechanisms of splicing," said James L. Manley, at Columbia University, NY. "With this kind of really rigorous genomic analysis,
I think these sequences will stand up and really be important elements."

However, Manley said he thought the paper had not addressed the issue of negative
splice regulators. "I think some elements they refer to as enhancer elements might
turn out to be negative or silencer elements," he said.

"The difference between fish and mammals is intriguing. It seems to give us some
insight into how splicing is done and what the rules are, what is allowed and what
is not," said Tomaso Poggio, professor in the Department of Brain and Cognitive Sciences, also at MIT. According
to Poggio, although the fish genome is much simpler, the greater diversity of regulatory
sequences might point to greater complexity at another level.

"Splicing is an important part of increasing diversity, and the estimates of genes
with alternative splicing keep going up," added Manley. "Splicing is becoming of more
and more interest as the number of genes in the human genome seems to keep going down."